Inhibitory effect of berberine on a nude mouse model of cutaneous squamous cell carcinoma and its mechanisms / 中华皮肤科杂志

ABSTRACT

Objective:To evaluate the effect of berberine on the volume and weight of, cell proliferation and apoptosis in transplanted cutaneous squamous cell carcinoma (cSCC) in nude mice inoculated with A431 cells, and to explore the possible mechanism underlying the inhibitory effect of berberine on cSCC.Methods:A431 cells were cultured, and A431 cell suspension was subcutaneously injected into the back of 20 nude mice to establish a nude mouse model of transplanted cSCC. On day 15 after inoculation, these tumor-bearing mice were randomly and equally divided into 4 groups low-, medium- and high-dose berberine groups intraperitoneally injected with 10, 20 and 25 mg/kg berberine solution respectively, and control group intraperitoneally injected with sodium chloride physiological solution. The treatment was performed once a day for 35 consecutive days. The tumor volume was measured before, and on days 7, 14, 21, 28 and 35 after the start of treatment. After the end of the experiment, the nude mice were sacrificed, and the tumors were removed and weighed to calculate the tumor growth inhibition rate. Histopathological examination was performed in these transplanted tumors, immunohistochemical study was conducted to determine the expression of Ki67, TUNNEL staining was conducted to determine the number of apoptotic cells in the transplanted tumor tissues, fluorescence-based quantitative PCR and Western blot analysis were employed to determine the mRNA and protein expression of apoptosis-related proteins Bax, Bcl-2, caspase-3 and Ezrin respectively. One-way analysis of variance was used for comparisons among multiple groups, and Dunnett- t test for comparisons of each berberine group with the control group. Results:Along with the increase in the dose of berberine and treatment duration, the tumor growth curve gradually became flat, the tumor growth was inhibited to different degrees in the berberine groups, and the high-dose berberine showed the strongest inhibitory effect on the tumor growth. The tumor growth inhibition rate was 31.05%, 66.68%, 76.49% in the low-, medium-, and high-dose berberine groups respectively, and the tumor weight was significantly lower in the 3 berberine groups than in the control group ( t = 4.07, 6.33, 7.26, respectively, all P < 0.01) . Along with the increase in the dose of berberine, histopathological examination of the transplanted tumors showed that the extent and area of tumor cell necrosis increased, while immunohistochemical study showed that the number of Ki67-positive cells gradually decreased. Moreover, the number of Ki67-positive cells was significantly lower in the low-, medium- and high-dose berberine groups than in the control group (all P < 0.01) , but the number of apoptotic cells was significantly higher in the berberine groups than in the control group ( P < 0.05 or < 0.01) . The mRNA and protein expression of Bax, Bcl-2, caspase-3 and Ezrin significantly differed among the 4 groups (all P < 0.01) . In addition, the mRNA expression of Bcl-2 was significantly lower in the low-, medium- and high-dose berberine groups than in the control group (all P < 0.01) , and the protein expression of Bcl-2 was significantly lower in the medium- and high-dose berberine groups than in the control group (both P < 0.01) ; Bax mRNA expression in the high-dose berberine group and caspase-3 mRNA expression in the medium- and high-dose berberine groups were significantly higher than the corresponding mRNAs in the control group respectively ( P < 0.05 or < 0.01) , and the protein expression of Bax and caspase-3 was significantly higher in the low-, medium- and high-dose berberine groups than in the control group (all P < 0.01) ; Ezrin mRNA expression was significantly higher in the high-dose berberine group than in the control group ( P < 0.01) , but its protein expression was significantly lower in the low-, medium-, and high-dose berberine groups than in the control group (all P < 0.01) . Conclusion:Berberine can inhibit the proliferation of A431 cells and promote their apoptosis in the transplanted cSCC of the nude mice, and then suppress the growth of transplanted cSCC in the nude mice to a certain extent, which may be related to the upregulation of Bax and caspase-3 and downregulation of Bcl-2 and Ezrin.